The overall objective of the proposed investigations is to elucidate the molecular mechanisms which regulate the quantities and types of glycerolipid synthesized in mammalian tissues. Studies on the regulation of microsomal enzymes synthesizing triacylglycerols, the principal storage fuel of animals, and of the microsomal enzymes synthesizing the glycerolphosphatides, ubiquitous components of biological membranes and of the serum lipoproteins are proposed using rat liver, isolated fat cells, and other rat tissues and in tissue culture cells. The approach enzymological. Previous work on the topography of triacylglycerol, phosphatidylcholine (PC), and phosphatidylethanolamine biosynthetic enzymes in the transverse plane of hepatic microsomal vesicles will be extended to enzymes involved in phosphatidylserine, phosphatidylinositol, and enzymes of ether lipid synthesis and of glycerolipid remodeling. Similar topographical studies will be performed in microsomes from other tissues. New methods of topographical analysis, including utilization of inside-out microsomes, and fusion of microsomes with phospholipid vesicles containing proteases, will be undertaken. A biochemical-genetic approach to regulation will be undertaken in Chinese hamster ovary cells (CHO) by testing whether LDL-PC will spare the choline requirements of these cells. The metabolism of LDL-PC will be studied to determine whether it is utilized intact or degraded in wild type, and mutant cell lines. Mutant strains defective in PC and glycerolipid metabolism will be selected and employed for regulatory studies. Solubilization and purification of microsomal glycerol-P and diacylglycerol acyltransferase will be undertaken. Regulatory investigations of glycerolipid biosynthetic enzymes in microsomes by phosphorylation, by phospholipid excesses, and by in vivo and in vitro modulation of phospholipid/protein ratios will be undertaken. The multifrontal attack on several important fronts should provide significant molecular insights about regulation of glycerolipid synthesis which is closely related to hyper- and hypo-lipoproteinemias, heart and vessel diseases, membrane and lipoprotein biogenesis, diabetes, cancer, obesity and the associated diseases of hypertention and stroke.